Uveal melanoma (UM) is the most common intraocular malignancy in adults and, despite successful local control, leads to substantial mortality due to early metastasis. Thus, there is an unmet need for novel systemic anticancer agents with activity against UM. Our preliminary studies identified distinct patterns of genotype-dependent melanoma cell sensitivity to targeted therapies. Cutaneous melanomas frequently harbor mutations in the BRAF gene, and are very sensitive to B-Raf inhibition, an approach that has been validated clinically and offers hope for patients with metastatic cutaneous melanoma. In sharp contrast, we demonstrated that uveal melanomas, that do not usually harbor BRAF mutations, are not sensitive to B-Raf inhibition. As a result, novel targeted therapies are urgently needed for UM. Somatic activating mutations in the genes encoding for the heterotrimeric G protein alpha subunits Gaq (GNAQ) and Ga11 (GNA11) are very frequently present in UM. Our preliminary data establish Gaq- induced signaling as the oncogenic driver in Gaq-mutant UM cells, via a pathway that involves PKCa, PKD and the steroid receptor coactivator (SRC)-3, a multifunctional oncogenic protein with potent growth promoting activity that, when overexpressed, confers poor prognosis in melanoma patients. We propose that pharmacological inhibition of Gaq or its downstream signaling cascade (PLCB, PKC, PKD and SRC-3) will exert anticancer activity in GNAQmt UM, thus representing a therapeutic opportunity for UM patients and a novel treatment paradigm for cancer. The overall goal of our proposal is to dissect the signaling pathway downstream of GNAQmt, the oncogenic driver in UM, and provide a roadmap for future studies that will lead to the development of personalized, targeted therapies. Our data highlight the role of the PLC/PKC/PKD/SRC3 signaling cascade in the GNAQmt pathway, providing insight into its cancer-promoting role and proof of concept that GNAQ-induced signaling can be selectively targeted in UM. We will explore further the indispensible role of SRC-3 in GNAQmt UM pathophysiology, by identifying its associated proteins and transcription factors by Mass Spectrometry. Using gene expression profiling analysis after siRNA- mediated silencing of these transcription factors, and chromatin immunoprecipitation (ChIP)- Sequencing (ChIP-Seq) analysis, we will determine their transcriptional target genes. This knowledge will help us identif and develop clinically applicable small molecule inhibitors of Gq and its downstream signaling cascade as novel targeted personalized therapies for GNAQmt UM. We have already identified a strong portfolio of compounds that can target the Gaq pathway and we will prioritize them for further validation studies in vitro and in vivo, using established, well-characterized UM cell line, primary UM cultures and a xenograft model of GNAQmt UM. These studies will be performed at the VA Boston Healthcare system under the direction of Dr Vasiliki Poulaki, an expert vitreoretinal surgeon as well as experienced molecular biology researcher, and in collaboration with Dr Bert W. O'Malley (Baylor College of Medicine), a world-renowned expert in SRC- 3. Our long term goal is to develop novel therapeutics targeting Gaq/SRC-3 in UM, with particular emphasis on rationally-designed combinational approaches based on a backbone of Gaq inhibition. As SRC-3 is overexpressed in a wide spectrum of malignancies, including breast, prostate, colon, lung and ovarian carcinomas, the identification of novel agents targeting SRC-3 will have wide therapeutic implications for other cancers as well. Dr Poulaki's goal is to develop safer and more effective therapies for UM, and to establish a Center of Excellence in the diagnosis and management of UM for Veteran patients across not only the East coast but across the country as well.